title: Alcohol dehydrogenase and cytochrome P450 2E1 can be induced by long-term exposure to ethanol in cultured liver HEP-G2 cells.
abstract: It has been shown in previous studies that liver HEP-G2 cells (human hepatocellular carcinoma) lose their ability to express active alcohol dehydrogenase (ADH) and cytochrome P450 2E1 (CYP2E1). Although both are ethanol-inducible enzymes, short-term exposure to ethanol does not cause any changes in expression or activity in cultured HEP-G2 cells. Therefore, we tested the effect of long-term exposure to ethanol on the expression and activity of both ADH and CYP2E1 in these cells. The expression of ADH and CYP2E1 was assessed at the mRNA and/or protein level using real-time PCR and Western blot analysis. Specific colorimetric assays were used for the measurement of ADH and CYP2E1 enzymatic activities. Caco-2 cells (active CYP2E1 and inactive ADH) were used as control cells. Significantly increased protein expression of ADH (about 2.5-fold) as well as CYP2E1 (about 1.6-fold) was found in HEP-G2 cells after long-term (12 mo) exposure to ethanol. The activity of ADH and CYP2E1 was also significantly increased from 12 ± 3 and 6 ± 1 nmol/h/mg of total protein to 191 ± 9 and 57 ± 9 nmol/h/mg of total protein, respectively. We suggest that the loss of activity of ethanol-metabolizing enzymes in cultured HEP-G2 cells is reversible and can be induced by prolonged exposure to ethanol. We are therefore able to reactivate HEP-G2 cells metabolic functions concerning ethanol oxidation just by modification of in vitro culture conditions without necessity of transfection with its side effect - enzyme overexpression.
authors.affiliation: Department of Cell and Molecular Biology and Center for Research of Diabetes, Metabolism and Nutrition, Third Faculty of Medicine, Charles University in Prague, Ruska 87, 100 00, Prague 10, Czech Republic, kamilabalusikova@seznam.cz.
authors.forename: Kamila
authors.initials: K
authors.lastname: Balusikova
authors.forename: Jan
authors.initials: J
authors.lastname: Kovar
mesh_terms: D000426:Alcohol Dehydrogenase / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D006528:Carcinoma, Hepatocellular / Q000201:enzymology* / Q000473:pathology; D019392:Cytochrome P-450 CYP2E1 / Q000096:biosynthesis* / Q000235:genetics; D004790:Enzyme Induction / Q000187:drug effects; D000431:Ethanol / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D056945:Hep G2 Cells; D006801:Humans; D008099:Liver / Q000187:drug effects / Q000201:enzymology; D008113:Liver Neoplasms / Q000201:enzymology* / Q000473:pathology; D010084:Oxidation-Reduction / Q000187:drug effects
chemical_list: D000431:Ethanol; D000426:Alcohol Dehydrogenase; D019392:Cytochrome P-450 CYP2E1